A fundamental question in development is how cells are specified. Elucidating mechanisms of cell-type specification is essential to the understanding and eventually treating human diseases, such as cancer, where this process is altered. One mechanism that specifies cell type is Asymmetric Cell Division, where a cell divides to produce two daughter cells that adopt distinct fates. Both intracellular asymmetric molecules and cell signaling appear to polarize neuroblasts, but a connection between the two mechanisms has been lacking. We have found that signaling is essential for certain asymmetric divisions that produce apoptotic cells and neural precursors and have identified molecules that are asymmetrically distributed in these divisions. The overall goal of our research is to understand how both cell intrinsic mechanisms and signaling regulate the fates of the neurons that are generated during these divisions. The proposal has three aims. 1) We propose to determine how the cytohesin GRP-1 regulates asymmetric cell divisions. Our model proposes that GRP-1 functions in V5 to regulate signaling between V5 and asymmetrically dividing neuroblasts, and we will test this idea rigorously for several divisions that require GRP-1 function. We will test the model that PI3K regulates GRP-1 and mediates its effects though ARF-1 and ARF-6. GRP-1's surprising localization to the nucleus raises the interesting possibility that cytohesins may function there, and we will ask whether GRP-1 carries out its functions in the nucleus. Finally, we will pursue RNG-1 and CNT-2, two molecules that may function in the GRP-1 pathway. 2) We propose to determine how Wnts regulate asymmetric divisions of the Q neuroblast, which also require both PIG-1 and GRP-1 function. Our preliminary experiments indicate that three Wnts and two Frizzled receptors are involved in these divisions. We will test the roles of the remaining Wnts and Wnt receptors, characterize the effects of these mutations on the divisions in more detail and test whether Wnt signaling acts in the GRP-1 or PIG-1 pathways. 3) Our analysis of GRP-1 and Wnt signaling indicates that cell signaling regulates asymmetric cell divisions. We will continue our EMS screens and begin RNAi screens to identify molecules that function in these signaling events, as well as molecules that act with HAM-1 and PIG-1.